Viral Vector Delivery of Gas Vesicle Genes
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Solution Overview
Problem
The use of gas vesicles (GVs) in mammalian cells is limited to transfection-compatible cell lines due to their large gene cluster size, restricting their application in broader biological contexts such as primary cells and endogenous tissues, and there is a need for an efficient and versatile platform to deliver these genes both in vitro and in vivo.
Innovation Solution
Viral vector compositions comprising one or more promoters operably connected to gas vesicle polynucleotides encoding GVA and GVS proteins, allowing for the formation of gas vesicles upon expression in cells, enabling efficient delivery and integration of acoustic reporter genes into mammalian cells and tissues.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If gas vesicle genes are delivered using traditional transfection methods, then gene expression can be achieved in transfection-compatible cell lines, but the application is limited to specific cell lines and requires tedious clonal expansions
Solution Approach 1:
The patent uses viral vectors as intermediary carriers to deliver gas vesicle genes to mammalian cells. The viral vector system mediates the transfer of genetic material from external sources into host cells, enabling gene delivery in cell types that are not amenable to traditional transfection methods. This intermediary approach bypasses the limitations of direct transfection while maintaining the ability to express gas vesicle genes across diverse cell types including primary cells and in vivo tissues.
2Quantity of substance
If the complete gas vesicle gene cluster is delivered as a single unit, then all necessary genes are provided, but the large size prevents efficient delivery to many cell types
Solution Approach 1:
The patent divides the complete gas vesicle gene cluster into multiple separate polynucleotide constructs, each containing one or more individual genes or gene subsets. These segmented polynucleotides are then delivered using multiple viral vectors in combination. This segmentation allows each vector to carry a manageable payload while collectively delivering the complete gene cluster, thereby improving delivery efficiency across different cell types while maintaining genetic completeness.
Solution Approach 2:
The patent combines multiple viral vectors, each carrying segmented polynucleotide constructs, to achieve complete gene delivery. The vectors work synergistically together, with each vector contributing a portion of the total genetic material. This merging approach allows the system to overcome the size limitations of individual vectors while still delivering the complete functional gene cluster required for gas vesicle formation.
3Measurement precision
If gas vesicles are used for acoustic imaging, then deep tissue imaging with high resolution is achieved, but the large gene cluster size restricts use to transfection-compatible cell lines
Solution Approach 1:
The patent employs viral vectors as intermediary delivery systems to overcome the limitations of traditional transfection methods. This mediation enables the delivery of gas vesicle genes to a broader range of biological contexts including primary cells, in vivo tissues, and cell types that were previously inaccessible, thereby expanding the versatility of acoustic imaging applications while maintaining high measurement precision.
Solution Approach 2:
By segmenting the gas vesicle gene cluster into deliverable polynucleotide constructs carried by viral vectors, the patent enables expansion into broader biological contexts. The segmented approach allows efficient delivery to primary cells and in vivo tissues that cannot accommodate large plasmid constructs through traditional transfection, thus expanding the range of applicable biological systems while preserving the acoustic imaging capabilities.
Data Source
AI summary
Disclosed herein include methods, compositions, and kits suitable for use in imaging of in situ gene expression. There are provided, in some embodiments, viral vector compositions. Disclosed herein includes a single viral vector comprising one or more gas vesicle assembly (GVA) gene(s) encoding one or more GVA protein(s), and one or more gas vesicle structural (GVS) gene(s) encoding one or more GVS protein(s). The one or more GVA protein(s) and the one or more GVS protein(s) can be capable of forming gas vesicles (GVs) upon expression in a cell.